Navegando por Palavras-chave "expression"

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    Chronic amphetamine transforms the emotional significance of a novel but not a familiar environment: implications for addiction
    (Cambridge Univ Press, 2011-08-01) Fukushiro, Daniela Fukue [UNIFESP]; Frussa Filho, Roberto [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)
    Both drug-induced locomotor sensitization and reactivity to novelty in rodents have been related to drug-craving mechanisms in humans. We investigated whether the exposure to a completely novel environment would modulate the expression of locomotor sensitization induced by repeated administration of amphetamine (Amp) in mice. in addition to locomotion, different open-field behavioural parameters were used to evaluate the possible involvement of anxiogenic-like effects induced by Amp, novelty or a combination of the two. in order to avoid misinterpretations due to different locomotor baseline conditions, we used an open-field illumination condition in which previous exposure to the apparatus did not modify locomotion (although it reliably increased grooming behaviour). Acute Amp administration increased locomotion in mice previously habituated to the open field (Hab) but not in mice exposed to the apparatus for the first time (Nov). This absence of Amp-induced locomotor activation in Nov mice may be related to higher anxiety-like levels, because these animals displayed longer freezing duration. However, only Nov mice developed locomotor sensitization. Because Amp challenge in Amp pre-treated Nov mice did not induce an increase in freezing behaviour, the locomotor sensitization in Nov mice might be related to the tolerance of Amp-induced anxiogenic-like behaviour in novel environments. Repeated Amp administration increased motivation to explore the environment in Nov mice in that these animals presented a within-session locomotion-habituation deficit. Our data suggest that a complex and plastic interaction between the anxiogenic and motivational properties of both novelty and Amp can critically modify the behavioural expression of craving-related mechanisms.
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    Cloning, expression and characterization of Bauhinia variegata trypsin inhibitor BvTI
    (Walter de Gruyter & Co, 2005-11-01) Souza, A. F. de; Torquato, RJS; Tanaka, A. S.; Sampaio, CAM; Universidade Federal de São Paulo (UNIFESP)
    A Bauhinia variegata trypsin inhibitor (BvTI) cDNA fragment was cloned into the pCANTAB5E phagemid. the clone pAS 1.1.3 presented a cDNA fragment of 733 bp, including the coding region for a mature BvTI protein comprising 175 amino acid residues. the deduced amino acid sequence for BvTI confirmed it as a member of the Kunitz-type plant serine proteinase inhibitor family. the BvTI cDNA fragment encoding the mature form was cloned into the expression vector, pET-14b, and expressed in E coli BL2I (DE3) pLysS in an active form. in addition, a BvTI mutant form, r(mut)BvTI, with a Pro residue as the fifth amino acid in place of Leu, was produced. the recombinant proteins, rBvTI and r(mut)BvTI, were purified on a trypsin-Sepharose column, yielding 29 and 1.44 mg/I of active protein, respectively, and showed protein bands of approximately 21.5 kDa by SDS-PAGE. Trypsin inhibition activity was comparable for rBvTI (K-I=4 nm) and r(mut)BvTI (K-i=6 nm). Our data suggest that the Leu to Pro substitution at the fifth amino-terminal residue was not crucial for proteinase inhibition.
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    Cysteine protease isoforms from Trypanosoma cruzi, cruzipain 2 and cruzain, present different substrate preference and susceptibility to inhibitors
    (Elsevier B.V., 2001-04-25) Lima, APCA; Reis, FCG dos; Serveau, C.; Lalmanach, G.; Juliano, L.; Menard, R.; Vernet, T.; Thomas, D. Y.; Storer, A. C.; Scharfstein, J.; Universidade Federal do Rio de Janeiro (UFRJ); Univ Tours; Universidade Federal de São Paulo (UNIFESP); Natl Res Council Canada
    Cysteine-proteinases from parasitic protozoa have been recently characterized as factors of virulence and pathogenicity in several human and veterinary diseases. in Chagas' disease, the chronic infection caused by Trypanosoma cruzi, structure-functional studies on cysteine proteases were thus far limited to the parasite's major isoform, a cathepsin L-like lysosomal protease designated as cruzipain, cruzain or GP57/51. Encoded: by a large gene family, cruzipain is efficiently targeted by synthetic inhibitors, which prevent parasite intracellular growth and differentiation. We have previously demonstrated that the multicopy cruzipain gene family includes polymorphic sequences, which could encode functionally different isoforms. We report here a comparative kinetic study between cruzain, the archetype of the cruzipain family, and an isoform, termed cruzipain 2, which is expressed preferentially by the mammalian stages of T. cruzi. Heterologous expression of the catalytic domain of cruzipain 2 in Saccharomyces cerevisae yielded an enzyme that differs markedly from cruzain with respect to pH stability, substrate specificity and sensitivity to inhibition by natural and synthetic inhibitors of cysteine proteases. We suggest that the structural-functional diversification imparted by genetic polymorphism or cruzipain genes may have contributed to T. cruzi adaptation to vertebrate hosts. (C) 2001 Elsevier Science B.V. All rights reserved.
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    A novel protein phosphatase 2A (MA) is involved in the transformation of human protozoan parasite Trypanosoma cruzi
    (Portland Press, 2003-09-14) Gonzalez, Jorge; Cornejo, Alberto; Santos, Marcia Regina Machado [UNIFESP]; Cordero, Esteban Mauricio [UNIFESP]; Gutierrez, Bessy; Porcile, Patricio; Mortara, Renato Arruda [UNIFESP]; Sagua, Hernan; Silveira, José F. da [UNIFESP]; Araya, Jorge E.; Univ Antofagasta; Universidade Federal de São Paulo (UNIFESP)
    Here we provide evidence for a critical role of PP2As (protein phosphatase 2As) in the transformation of Trypanosoma cruzi. in axenic medium at pH 5.0, trypomastigotes rapidly transform into amastigotes, a process blocked by okadaic acid, a potent PP2A inhibitor, at concentrations as low as 0.1 muM. 1-Norokadaone, an inactive okadaic acid analogue, did not affect the transformation. Electron microscopy studies indicated that okadaic acid-treated trypomastigotes had not undergone ultrastructural modifications, reinforcing the idea that PP2A inhibits transformation. Using a microcystin-Sepharose affinity column we purified the native T cruzi PP2A. the enzyme displayed activity against 12 P-labelled phosphorylase a that was inhibited in a dose-dependent manner by okadaic acid. the protein was also submitted to MS and, from the peptides obtained, degenerate primers were used to clone a novel T cruzi PP2A enzyme by PCR. the isolated gene encodes a protein of 303 amino acids, termed TcPP2A, which displayed a high degree of homology (86%) with the catalytic subunit of Trypanosoma brucei PP2A. Northern-blot analysis revealed the presence of a major 2.1-kb mRNA hybridizing in all T cruzi developmental stages. Southern-blot analysis suggested that the TcPP2A gene is present in low copy number in the T cruzi genome. These results are consistent with the mapping of PP2A genes in two chromosomal bands by pulsed-field gel electrophoresis and chromoblot hybridization. Our studies suggest that in T cruzi PP2A is important for the complete transformation of trypomastigotes into amastigotes during the life cycle of this protozoan parasite.